Single cell, whole genome analysis of the aging human cardiomyocyte

衰老人类心肌细胞的单细胞、全基因组分析

• 批准号: 10326403
• 负责人: Sangita Choudhury
• 金额: $ 87.2万
• 依托单位: BOSTON CHILDREN'S HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-01-15 至 2025-12-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10326403
• 关键词: Adopted; Adult; Age; Aging; Algorithms; Alleles; Autopsy; Brain; Cardiac; Cardiac Myocytes; Cardiomyopathies; Cardiovascular Diseases; Cardiovascular Pathology; Cell Nucleus; Cell Respiration; Cells; Copy Number Polymorphism; DNA; DNA Damage; DNA Sequence Alteration; DNA sequencing; Data; Development; Disease; Disease Progression; Dropout; Elderly; Environmental Risk Factor; Enzyme-Linked Immunosorbent Assay; Freezing; Genetic; Genome; Genomic DNA; Genomics; Goals; Heart; Heart Diseases; Heart failure; Human; In Vitro; Individual; Inherited; Interphase Cell; Knowledge; Lead; Link; Longitudinal Studies; Malignant Neoplasms; Measures; Methods; Modeling; Mutagens; Mutation; Myocardial dysfunction; Myocardium; Neurons; Newborn Infant; Organ; Oxidative Stress; Pathologic; Patients; Pattern; Phenotype; Ploidies; Point Mutation; Premature aging syndrome; Process; Radiation; Radiation therapy; Research; Resolution; Risk Factors; Role; Sampling; Series; Single Nucleotide Polymorphism; Somatic Mutation; Techniques; Testing; Time; Troponin T; Variant; age related; aged; base; cancer radiation therapy; cardiovascular risk factor; cell type; disorder risk; epidemiology study; genome analysis; genome sequencing; genome-wide; genotoxicity; human old age (65+); innovation; insight; novel; prevent; radiation during childhood; radiation effect; whole genome

Project Summary/Abstract Recent evidence suggests that environmental factors causing somatic mutations during the lifetime have a more crucial role, not only in cancer but also in other common diseases, including heart failure. Recent studies have also shown that somatic single nucleotide variants (sSNV) accumulate even in nondividing cells, such as neurons in the human cortex, resulting in thousands of sSNV per neuronal genome by old age. However, genomic DNA changes in aging cardiomyocytes (CM) remain poorly understood. The accumulation of somatic DNA mutations over time has recently been demonstrated to be a hallmark of aging in many human cell types. The current study aims to determine the landscape and role of somatic mutations in aging and cardiac disease by adopting a new technique that allow deep whole-genome sequencing of DNA isolated from single CM taken from the frozen postmortem heart. The first Aim of this study is to evaluate the somatic mutational burden (sSNVs) in aging CM genome. We will also compare CM mutational burden with postmitotic cells from another organ (neurons) to define differences in accumulation rate during aging. In the second Aim, we will ask what are the mutational signature and the mechanisms of mutation formation in the aging human heart and if the heart mutational signature is different than the brain mutational signature. Further to recapitulate the mutational signature and related phenotype in the heart we will directly induce oxidative stress in an in vitro culture model of primary CMs. The final Aim will focus on evaluating the genotoxic effect of radiation in CMs after childhood radiation therapy and the role of radiation in premature aging. The proposed research is significant for the comprehensive, results-based development of strategies for understanding natural aging and disease progression in the human heart. Together with the planned characterization of mutational signatures, the anticipated results may provide knowledge to develop new strategies for preventing the heart disease associated with aging. The proposed study is only possible because of a series of innovations that are, at this time, uniquely available to our research team, 1) a novel method to isolate single CM nuclei from frozen myocardium based on CM ploidy and nuclei cardiac troponin T expression. 2) A major breakthrough by developing “LiRA” and “PhaseDel” algorithm to call sSNV and sSV confidently from tetraploid cells that considers cell-specific depth distributions of DNA sequencing and allele-dropout rates in scWGS data. For the first time, our study will reveal the landscape of somatic mutations, genomic changes during aging and after radiation therapy in human heart muscle cells in a single-cell resolution. In the long term, this study will provide insights that might allow blocking some of the mutational processes ameliorating age-related myocardial dysfunction.

项目摘要/摘要 最近的证据表明，在生命中引起躯体突变的环境因素具有 不仅在癌症中，而且在其他常见疾病中，包括心力衰竭，更关键的作用。最近的研究 还表明，体细胞单核苷酸变体（SSNV）即使在非各个细胞中也积累 人皮层中的神经元，到老年，每神经元基因组成千上万的SSNV。然而， 衰老心肌细胞（CM）的基因组DNA变化仍然很了解。躯体的积累 随着时间的流逝，DNA突变最近被证明是许多人类细胞类型中衰老的标志。 当前的研究旨在确定体细胞突变在衰老和心脏疾病中的作用 通过采用一种新技术，该技术允许从单个CM中分离出的DNA的深度全基因组测序 从冷冻的验尸心脏中。这项研究的第一个目的是评估躯体突变伯恩 （SSNV）在衰老CM基因组中。我们还将比较CM突变烧伤与另一个的有丝分裂细胞 器官（神经元）定义衰老过程中累积率的差异。在第二个目标中，我们会问什么 是突变特征和人类衰老的突变形成的机制，以及是否 心脏突变特征与大脑突变特征不同。进一步概括突变 签名和相关表型在心脏中我们将直接诱导体外培养模型中的氧化应激 主要CMS。最终目的将重点放在评估儿童之后CMS辐射的遗传毒性作用上 辐射疗法和辐射在过早衰老中的作用。拟议的研究对 全面的，基于结果的理解自然衰老和疾病的策略 人心的进展。加上突变特征的计划表征， 预期的结果可能会提供知识，以制定预防心脏病的新策略 与衰老有关。拟议的研究仅是可能的，因为这是一系列创新 时间，我们的研究团队独特地使用，1）一种新的方法，可以将单个CM核心与冷冻隔离 基于CM倍性和核心脏肌钙蛋白T表达的心肌。 2）一个重大突破 开发“里拉”和“ phasedel”算法，以从四倍体细胞中自信地调用SSNV和SSV 考虑了SCWGS数据中DNA测序和等位基因降低率的细胞特异性深度分布。为了 第一次，我们的研究将揭示躯体突变的景观，衰老期间的基因组变化 单细胞分辨率中人类心肌细胞中的放射治疗。从长远来看，这项研究将 提供可能允许阻止一些与年龄相关的突变过程的见解 心肌功能障碍。